Originally regarded as intermediates in lipid biosynthesis (Kent, Anal. Rev. Biochem. 64:315-343, 1995), phosphatidic acid (PA) and one of its precursors, lysophosphatidic acid (LPA), have also been identified as phospholipid signaling molecules that affect a wide range of biological responses (McPhail et al., Proc. Natl. Acad. Sci. USA 92:7931-7935, 1995; Williger et al., J. Biol. Chem. 270:29656-29659, 1995; Moolenaar, Curr. Opin. Cell Biol. 7:203-210, 1995).
Cellular activation in monocytic and lymphoid cells is associated with rapid upregulation of synthesis of phospholipids (PL) that includes phosphatidic acid (PA), diacylglycerol (DAG) and glycan phosphatidylinositol (PI). Phosphatidic acids (PA) are a molecularly diverse group of phospholipid second messengers coupled to cellular activation and mitogenesis (Singer et al., Exp. Opin. Invest. Drugs 3:631-643, 1994). Compounds that would block PA generation and hence diminish the signal involved in cell activation may therefore be of therapeutic interest in the area of inflammation and oncology. Lysofylline (1-(R)-(5-hydroxyhexyl)-3,7-dimethylxanthine) (Singer et al., Exp. Opin. Invest. Drugs 3:631-643, 1994; and Rice et al., Proc. Natl. Acad. Sci. USA 91:3857-3861, 1994) has been found to be an effective inhibitor of cellular activation by blocking the synthesis of a specific phosphatidic acid (PA) species produced by lysophosphatidic acid acyltransferase (LPAAT) in activated monocytic cells (Rice et al., Proc. Natl. Acad. Sci. USA 91:3857-3861, 1994). PA can be generated through hydrolysis of phosphatidycholine (PC) (Exton, Biochim. Biophys. Acta 1212:26-42, 1994) or glycan PI (Eardley et al., Science 251:78-81, 1991; Merida et al., DNA Cell Biol. 12:473-479, 1993), through phosphorylation of DAG by DAG kinase (Kanoh et al., Trends Biochem. Sci. 15:47-50, 1990) or through acylation of LPA at the SN2 position (Bursten et al., Am. J. Physiol. 266:C1093-C1104, 1994). Compounds that would block PA generation and hence diminish lipid biosynthesis and the signal involved in cell activation may therefore be of therapeutic interest in the area of inflammation and oncology as well as obesity treatment.
The genes coding for LPAAT have been isolated in bacteria (Coleman, Mol. Gen. Genet. 232:295-303, 1992), in yeast (Nagiec et al., J. Biol. Chem. 268:22156-22163, 1993) and in plants (Brown et al., Plant Mol. Biol. 26:211-223, 1994; and Hanke et al., Eur J. Biochem. 232:806-810, 1995) using genetic complementation techniques. The cloning of a mammalian version of LPAAT has not been reported. Homology among the bacterial, yeast and plant LPAAT is only found in a very few block of three or at most four amino acids scattered throughout the sequences (Brown et al., Plant Mol. Biol. 26:211-223, 1994). Further, there is a need in the art for recombinant LPAAT from a mammalian source to enable compound screening for LPAAT inhibitors for the development of specific compounds that would inhibit this enzyme.